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Genetic engineering of virus-specific T cells with T-cell receptors recognizing minor histocompatibility antigens for clinical application

¹ Laboratory of Experimental Hematology, Leiden University Medical Center, Leiden, The Netherlands
² ProImmune Ltd., The Magdalen Centre, Oxford Science Park, Oxford, UK

Correspondence: Marieke Griffioen, Department of Hematology, C2-R, Leiden University Medical Center, P.O. Box 9600, 2300 RC Leiden, The Netherlands. E-mail:m.griffioen{at}lumc.nl

Background: Donor lymphocyte infusion is an effective form of adoptive immunotherapy for hematologic malignancies after allogeneic stem cell transplantation. Graft-versus-host disease, however, often develops due to recognition of ubiquitously-expressed minor histocompatibility antigens. Transfer of T-cell receptors recognizing hematopoiesis-restricted minor histocompatibility antigens to virus-specific T cells may be a powerful anti-tumor therapy with a low risk of graft-versus-host disease. The purpose of this study was to develop an optimal T-cell receptors-encoding multi-cistronic retroviral vector and an efficient method for generating T-cell receptors-engineered virus-specific T cells.

Design and Methods: Retroviral vectors encoding the T-cell receptors for the hematopoiesis-restricted minor histocompatibility antigen HA-2 with and without selection markers were compared for T-cell receptors surface expression and HA-2-specific lysis. In addition, two different methods, i.e. peptide stimulation of CD8⁺ cells and Pro5® MHC pentamer-based isolation of antigen-specific T cells, were investigated for their efficiency to generate T-cell receptors-transduced virus-specific T cells.

Results: Bi-cistronic vectors without selection markers most efficiently mediated T-cell receptors surface expression and HA-2-specific lysis. Furthermore, both methods were useful for generating gene-modified cells, but the purity of virus-specific T cells was higher after pentamer isolation. Finally, the capacity of gene-modified cells to express the transgenic T-cell receptors at the cell surface markedly differed between virus-specific T cells and was correlated with lysis of relevant target cells.

Conclusions: Our data support T-cell receptors gene transfer to pentamer-isolated virus-specific T cells using bi-cistronic retroviral vectors and illustrate the relevance of selection of gene-modified T cells with appropriate transgenic T-cell receptors surface expression for clinical gene therapy.

Key words: allogeneic stem cell transplantation, immunotherapy, gene therapy, T cell receptor.